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The Diverse Applications of
Mass Spectrometry
Contents
How Mass Spectrometry Has Changed
the Cancer Diagnostics Game
How to Test for Heavy Metals in Cannabis
Steve Pennington on 10 Years of
Proteomic Progress
Rapid Detection of Designer Drugs
Presents Challenges for Law Enforcement
and Forensic Toxicology 10
How Advances in Mass Spectrometry Are
Helping the Food and Beverage Industry
Measuring How the Brain Metabolizes
in Real-time
Mass Spectrometry’s Important Role in
Identifying Explosives in the Environment 17
Mass Spectrometry in
Biopharmaceutical Discovery
Helping Analytical Chemistry Embrace Big
Data23
Developing FTICR Mass Spectrometry
Instruments with Unique Capabilities
Foreword
In today’s world, the diversity of mass
spectrometry instrumentation and applications
thereof is simply mind-boggling. The mid to late
19th century heralded the emergence of the
concept that later led to the development of mass
spectrometers that could physically ‘weigh’ the
mass of a molecule. Over the last 150 years, this
analytical instrumentation has developed into
the fascinating and remarkably diverse field of
mass spectrometry (MS) with practical, modern-
day applications covering a myriad of topics.
MS is used in forensics and anti-terrorism to
analyze chemical warfare agents and explosives.
4 These not only pose immediate danger to
the safety and well-being of people; but can
6 have long-term hazardous implications for
humankind, wildlife and the ecosystem. MS
also aids in monitoring drug abuse by assisting
8 law enforcement in the rapid detection and
identification of designer drugs and narcotics;
and plays a key role in maintaining societal
health by evaluating and ensuring the safety of
food and beverages. Similarly, in the medicinal
plant industry, heavy metal levels in cannabis
are monitored by MS to guarantee the continued
well-being of patients.
12
MS furthers our access to new, innovative
therapeutic approaches to diagnose, monitor
and treat disease. Applications include real-time
15 monitoring of brain cell metabolism in animals for
toxicological profiling and drug discovery. Other
advances in MS not only provide information on
higher-order protein structure, contaminating
proteins and the in vivo behavior of emerging
biotherapeutics; but also, on the efficacy and
20 safety of these drugs. Furthermore, MS is used
to understand aberrant signaling pathways
for drug development, or to identify molecular
signatures from the circulating proteome of
a patient that can be exploited to overcome
treatment resistance and relapse. Indeed, MS
can now even guide surgeons in real-time to
25 precisely resect cancerous tissue.
This eBook highlights the broad spectrum
of applications that integrally utilize mass
spectrometry. Such a powerful technique now
spans diverse fields and has an extensive and
far-reaching impact on society.
 The Diverse Applications of Mass Spectrometry

How Mass
MasSpec Pen Detects Cancer in Seconds and
One-ups the iKnife


Spectrometry A hand-held device that connects to a mass spectrometer and

identifies cancerous tissue during surgery hit the headlines last

Has Changed year, claiming to deliver results in ten seconds. Termed MasSpec

Pen, the device contains a probe that can soak up molecules
from human tissue and send to a mass spectrometer wheeled

the Cancer into the surgery room. Within a matter of seconds, molecular
analysis by an ambient ionization MS technique indicates which

Diagnostics areas of tissue are cancerous and should be removed by surgery.

The device can also sample tissue non-destructively, offering an

Game
advantage over a similar device called iKnife that causes tissue
damage due to electrocauterization.



Charya Wickremasinghe, PhD In a test of 253 patients with lung, breast, thyroid or ovarian
cancer, the MasSpec Pen was able to provide a tissue diagnosis
with over 96 percent accuracy.  Even in marginal regions that

W
ith a history that spans one hundred years, mass contained mixed tissue and cellular compositions, the device was
spectrometry (MS) is one of the most widely used able to detect cancer. 


analytical techniques in bioscience and medical
research. Since the development of the first modern mass During development, researchers used a benchtop liquid
spectrometer in 19181, the technique has advanced steadily over chromatography-tandem mass spectrometry (LC-MS/MS)
time and found its way into a range of applications from forensic system with quadruple precursor ion selection, and high-
toxicology to cancer diagnostics. Bulky, expensive equipment resolution, accurate-mass (HRAM) Orbitrap detection.2 Since
that defined historical mass spectrometers are less common this is a large instrument that takes up considerable space in
now, and have been replaced by faster, more sensitive, and the surgery room, developers are already looking into getting
cost-effective workflows. Newer spectrometers are also largely reliable data from a more compact spectrometer.   
automated and user-friendly, making them more adaptable for
clinical use.

 Using Quantitative MS to Help Guide Lung Cancer
Therapy

Although histopathology is still the gold standard for diagnosing
cancer, molecular analysis using MS has gained traction in the A team of researchers at the Virginia Commonwealth
recent years for detecting tumors, monitoring progression, and University lead by Professors Adam Hawkridge (Department
even predicting treatment response. 
 of Pharmaceutics, School of Pharmacy) and David Gewirtz
(Department of Pharmacology and Toxicology, School of

Technology Networks 2018 4 TechnologyNetworks.com

 The Diverse Applications of Mass Spectrometry
Medicine) are utilizing quantitative mass spectrometry to
uncover clues related to treatment resistance and relapse of non-
small cell lung cancer (NSCLC). 

 mass spectrometry into a machine learning platform for
“Mass spectrometry can be used on a number of levels to better
understand NSCLC signaling pathways for drug development,
or to identify molecular signatures that can be exploited to
overcome NSCLC treatment resistance,” says Hawkridge. “For
example, in our most recent LC-MS/MS study of treatment
resistance, we found that senescence – rather than autophagy
– associated proteins were more prominently secreted in the
treatment resistant-cells in response to ionizing radiation.”

 that are often missed.”
In a previous study by Hawkridge’s team, LC-MS/MS analysis
allowed the identification of 364 secreted proteins that could be
further studied for their potential diagnostic value as a function
of p53 expression, irradiation, and functional autophagy status
in the context of NSCLC.3  The lead author of the study, Dr.
Emmanuel Cudjoe, explained that cancer cells respond to stress
by activating a process called autophagy in which organelles are
recycled and energy production is maintained, leading to tumor
growth, therapeutic resistance and relapse. 


“Where autophagy contributes to treatment resistance, inhibition
can restore tumor sensitivity to chemo/radiotherapy. Our long-
term goal is to identify molecular signatures and pathways of
autophagy that can be targeted to improve the sensitivity to
radiotherapy,” Cudjoe adds. 

 proteins that are very close in mass/charge ratio. Röder’s team is
MS-based Test Helps to Select the Right Patients
for the Right Type of Immunotherapy
 cancer such as NSCLC.
For treating patients with metastatic melanoma, there are several
different types of immunotherapeutic regimens available,
including mono-therapies and combinations of immune-
checkpoint inhibitors. However, selecting and optimizing the
right treatment choice for each patient is challenging. To assist in
treatment selection, a new MS-based blood test was developed
by Biodesix, Inc. in collaboration with scientists from the NYU
Langone Medical Center, New York, Yale University School of
Medicine, New Haven, and the Massachusetts General Hospital,
Boston.

 incorporated into the proteomics databases for additional levels
“We developed an anti-programmed cell death protein-1 (PD-1)
melanoma blood test using a machine learning platform with
a combination of clinical outcome data and expression data
of circulating proteins based on mass spectral analysis,” says
Dr. Heinrich Röder, Chief Technology Officer at Biodesix.
“We’ve shown this test to predict survival in patients receiving
PD-1 inhibitors. It can also identify patients who are resistant
to monotherapy, but may in fact benefit from combination
immunotherapy instead.”


Technology Networks 2018
Scientists at Biodesix input data acquired from Matrix Assisted
Laser Desorption/Ionization Time-of-Flight (MALDI-ToF)
multivariate test design.4  “MALDI allows us to take a real-time
snapshot of a patient’s circulating proteome, which is derived
from the tumor, tumor microenvironment, and normal host
tissue. These proteins have direct regulatory effects on the
immune system, especially the interplay of the innate and
adaptive system,” Röder explains. “We do not need to choose
proteins based on pre-conceived notions of relevance, as one has
to do in fixed panel approaches, and hence can observe effects



Biodesix uses a special MALDI-ToF method for protein analysis,
termed Deep MALDI™, which is aimed at reducing the noise
in mass spectral data by increasing the number of laser shots.
“With this method, we can measure more proteins in a robust
and reproducible way with a high sample throughput and small
sample volume,” says Röder. “We also experience a much wider
coverage of reliably detectable proteins and peptides compared
to traditional MALDI-ToF.” 


Although Deep MALDI can function across an abundance
range of 4-5 orders of magnitude, it struggles to pick up very
low abundance proteins outside the range of the classical plasma
proteome. Other limitations of the method include reduced
sensitivity at masses higher than 30kDa, and low resolution of
currently working on improving this technology and expanding
it to investigate immunotherapy responses in other types of



Future of MS-based Proteomics

With the advent of increasingly sophisticated technologies,
MS instruments are becoming faster and more sensitive with
respect to molecular characterization. Hawkridge believes that
advances in analytical figures of merit will drive new molecular
insight in areas such as protein post-translational modifications
(e.g., glycosylation). He elaborates, “As genomic sequencing
becomes more prevalent, these genetic perturbations are being
of data analysis. That is just a small sampling of the untapped
potential of mass spectrometry-based cancer proteomics.”
References
1. A. J. Dempster, Phys. Rev. 11, 316–325. (1918)

2. J. Zhang et al., Sci. Transl. Med. 9, (2017)

3. E. K. Cudjoe et al., Cancer Res. 77, 227–227. (2017)

4. J. S. Weber et al., Cancer Immunol. 6, 79–86. (2018)
5 TechnologyNetworks.com
 The Diverse Applications of Mass Spectrometry
How to Test for
Heavy Metals in
Cannabis
Mike May, PhD
C
annabis collects heavy metals. It “absorbs heavy metals
from the soil, water and air,” says Bob Clifford, general
manager at Shimadzu Scientific Instruments. These
elements—arsenic, cadmium, lead, mercury and others—can
trigger a range of health problems, including cancer and heart
disease. Although cannabis for human consumption or smoking
should be as free from heavy metal contamination as possible,
that’s not always the case. When Sammy Hagar wrote “sparks fly
in the middle of the night,” little did he know that this line could
describe not just heavy-metal music, but the concerns of testing
of cannabis for these elements.
Today’s ‘sparks’ fly around the need and necessary extent of
testing cannabis for heavy metals. “Heavy-metal contamination
will vary from farm to farm and different strains will have
different uptake rates of metals, thus the importance of testing,”
Clifford explains.
Regardless of where cannabis comes from, it can include heavy
metals. “All agricultural products, including cannabis, contain
some amount of heavy metals,” says GenTech Scientific. “The
use of things such as fertilizers and pesticides in agricultural
applications increase the abundance of these metals in soils and
water.”
Beyond coming from natural sources and agricultural methods,
heavy metals can infiltrate cannabis-based products in other
Technology Networks 2018
ways. As an example, GenTech Scientific notes: “Contamination
can also occur during the processing of cannabis.”
Despite the wide range of potential sources of heavy metals,
it is not inevitable that every cannabis sample includes these
elements—at least not at high levels. “With the correct
atmosphere, clean soil and water, and limited use of pesticides
and fertilizers, you can keep your levels below the acceptable
safe limit,” according to GenTech Scientific. Nonetheless, this
company adds, “It is important for growers to monitor the levels
to ensure their products are in compliance with their state’s
regulatory guidelines.” And those can vary considerably.
Where regulators do require testing cannabis for heavy metals,
the list usually includes arsenic, cadmium, lead and mercury.
“Other states, like Maryland, include additional elements to
be tested for such as barium, chromium, selenium and silver,”
Clifford says.
Testing Tools
Cannabis can be tested for heavy metals in many ways, such
as various forms of atomic spectrometry, including atomic
absorption (AA), inductively coupled plasma optical emission
spectroscopy (ICP-OES) and inductively coupled plasma mass
spectrometry (ICP-MS). “For AA, the method used would have
to be the more sensitive graphite furnace atomic absorption—
GFAA—since the flame AA method would not be sensitive
enough for most elements,” Clifford explains. “Also, mercury
must be measured by a method called cold vapor atomic
absorption spectroscopy—CVAAS—due to poor sensitivity by
AA.”
In general, flame techniques can measure elements at low parts
per million, and GFAA goes down to low parts per billion.
“Also, the AA method usually measures one element at a time,”
Clifford says. “ICP and ICP-MS are techniques capable of
measuring multiple elements simultaneously.” Nonetheless,
6 TechnologyNetworks.com
 The Diverse Applications of Mass Spectrometry

using ICP to test cannabis for heavy metals, often requires a way not test for other consumed products like cannabis, especially
to enhance its sensitivity, such as introducing the sample with an when involving immune-compromised patients?” Clifford asks.
ultrasonic nebulizer (USN). “The USN can increase sensitivity “There needs to be federal oversight of testing for heavy metals,
up to a factor of 10,” Clifford says. as well as pesticides, residual solvents, mycotoxins and other
contaminates so testing is harmonized throughout the United
From Aeos Labs, analytical chemist Anya Engen says, “ICP-MS States.”
offers the best sensitivity and is the method used in our lab.”
She adds, “The FDA and United States Pharmacopeia have When asked if any testing improvements would be useful, Engen
standardized methods for heavy-metals analysis, which are very says, “Measuring heavy-metal levels in the soil and water where
useful resources in the fledgling cannabis-testing industry, where cannabis is grown and correlating the data to cannabis-generated
regulation is slow to catch up.” data would be helpful in illustrating an overview of possible
contamination sources.” She adds, “Being in Hawaii with an
To test for arsenic, scientists often need special sample active volcano, the heavy metals produced and dispersed by the
preparation, such as liquid chromatography (LC), because volcano is a possible source of contamination in Hawaii-grown
arsenic can be in inorganic or organic forms. The inorganic cannabis.”
form is more toxic. “If one wants to test for the different forms,
LC-ICP-MS is utilized to separate and detect the different Without testing cannabis for heavy metals, sparks could fly from
compounds,” Clifford says. “States currently only require total more than guitars as customers insist increasingly on safe, tested
arsenic, so speciation of this compound is not required.” products.

Tomorrow’s Testing

Not all regulators even require heavy-metals testing for cannabis

at the moment. “We test for heavy metals in the USA’s food
supply system, enforced by FDA and USDA, so why would we

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Technology Networks 2018 7 TechnologyNetworks.com

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 The Diverse Applications of Mass Spectrometry
Steve
Pennington
on 10 Years
of Proteomic
Progress
Ruairi MacKenzie
S
teve Pennington, Professor of Proteomics at University
College Dublin, has spent over 35 years in research.
Focusing on using mass spectrometry to measure and
characterize proteins, Steve has made contributions not only in his
80+ published papers, but also as part of the editorial board of the
Journal of Proteomics and other journals, and as the lead organizer of
the Human Proteome Organization’s (HUPO) World Congress
held in Dublin in 2017. We asked Steve how the field of mass
spectrometry has advanced in the last decade of research.
Q: Just over ten years ago you co-authored a
paper entitled Guidelines for the Next 10 Years of
Proteomics. We’re just about 12 years on from that
paper - what predictions did you make back then
and how accurate were you?
A: I recently went back and had a careful re-read of the paper and
I think rather than attempting to predict the future of proteomics,
the paper was reporting a summary of where things had got up to,
Technology Networks 2018
as well as where some of the issues and limitations lay, and maybe
how those limitations could be addressed by producing some
guidelines for the next 10 years. It wasn’t really predicting what
might happen in the next 10 years.
And it’s been really interesting to look back to see what
has happened because I guess when you’re involved in the
developments day by day and always looking forward, it may
seem that not a lot is happening. So, when you get the opportunity
to review the last 10 years, it’s apparent that there have been
remarkable changes. Maybe not as remarkable as we would have
liked them to be, but still pretty significant.
I think one of the key things the paper was really trying to push
was the development of standards and guidelines for proteomics
that would allow greater reproducibility and greater confidence in
the data that was coming from proteomics studies. And in the last
10 years there’s been a huge effort in that area, such that we are now
at a position where, when proteomics experiments are designed and
done properly, which they are in a good proportion of the field, the
data that we get from them is really robust and reliable.
The Human Proteome Organization (HUPO) has been
instrumental in driving the developments of those guidelines.
That’s been supported, and in many instances, accelerated by some
of the key journals in the field. And so, we have reached the position
now where standards for the design and execution of experiments
and also, importantly, the analysis of the data have reached a stage
where they do provide reliable, reproducible data.
That may not sound like a major advance, but in terms of being
able to build from the proteomics discoveries that have been made,
and convert them into useful applications, it’s been fundamentally
important.
These developments have been happening in parallel with, I
suppose, a general concern about the reproducibility of biological
studies and cancer research. In the last 10 years and relatively
8 TechnologyNetworks.com
 The Diverse Applications of Mass Spectrometry
recently, some high profile publications have reported that fairly
large numbers of studies, particularly in the drug discovery arena,
have not been very reproducible. Notably, drug discovery and
development companies have found it difficult to reproduce studies
that are reported in the literature.
So, I think getting to reproducibility has been a very significant
step forward and for me one of the take-home messages from the
HUPO 2017 meeting was, that over the last 10 years the technology
has advanced, and as importantly, the reliability of the data that’s
being produced from the technology has also.
Q: I am regularly contacted about publishing
content on data integrity, for example, on a new
integrity pipeline or project. In that 10-year period,
what are the particular projects or advancements
that have improved data integrity within
proteomics?
A: I think if, as we now do, you have standards that need to be met
for publication of data, a checklist of the journal’s requirements for
submitting data, that has knock-on effects for the generation of the
data. The development of transparent databases, like the PRIDE
and PeptideAtlas databases which are large repositories of data, has
meant that others are readily able to reanalyze the raw data that’s
been produced in adherence to the standards.
I suppose it’s human nature - when you know that your data is
potentially going to be scrutinized by others, there becomes a
greater emphasis at home in making sure that that data is robust
and reliable.
Q: What about quantity, as well as quality? Have
technological advances had a big impact on the
amount of data produced?
A: Absolutely. So, 10 years ago, one of the bedrock technologies
of proteomics was the use of 2D gels, relatively slow-throughput
and producing modest amounts of data. This has largely been
superseded by doing LC/MS, both bottom-up and top-down,
but mainly bottom-up, where very complex protein mixtures are
digested into peptides and those peptides (10’s of 1,000’s from a
single sample) are analysed in the mass spectrometer. And so, there
are very, very large data sets.
In the last few years, there’s been the development of an approach
where the data is acquired in the mass spectrometer in a fully
comprehensive data-independent acquisition mode. So, whereas
in the past, the mass spectrometer would, in real time, make
some decisions about what peptides to fragment and get data
from, the current trend is to take every peptide that’s put into the
mass spectrometer and fragment it and acquire data from it, in a
completely unbiased, comprehensive manner.
Technology Networks 2018
Obviously, the data sets that come out of this approach are
significantly richer in terms of the amount of information produced.
They’re also more challenging in terms of the interpretation of the
data, but then great tools are being developed for analyzing these
complex data sets.
We’ve moved on from situations, I think, where if you put one
sample into the mass spectrometer and you did that at least
three times, you would get overlapping data but there would
be differences in the data obtained from the three separate
mass spectrometry runs, and that’s with large amounts of data.
Now, there’s greater confidence because you’re doing that
comprehensively and the same every time, so there’s a larger
amount of data and it’s almost completely overlapping between the
runs. And this comprehensive approach can be viewed, if you like,
almost as a permanent record of that sample, the proteome of the
sample.
Q: If you were going to speculatively write another
review looking forward 10 years, do you think you’d
be able to do that, or is the field moving too fast
now?
A: Yes, I think we could do it and I’m hoping to take that
opportunity during the HUPO Congress in 2018. I think it would
be really interesting and valuable to have a multi-authored article
that looks carefully at what we’ve achieved in the last 10 years and
what the future holds; current limitations and future opportunities.
I’m particularly excited about the future. I haven’t really touched on
it much, on some of the work that’s being done on taking the now
very comprehensive biomarker discovery data sets that have been
generated and converting those into multiplex targeted protein
assays with the aim that these will become the advanced diagnostic
tests of the future to support precision medicine.
We know that diseases are very heterogenous and complex and
that there’s a lot of patient heterogeneity, and so the ability in the
future to be able, in an analytical robust way, measure multiple
(100’s of) proteins, means that we have a great opportunity to
develop new, more sophisticated diagnostic tests. More importantly,
it is possible that these new diagnostic tests will exploit the
developments in targeted mass spectrometry on triple quadrupole
mass spectrometers that have been made over the last several
years. Together, new tests and instrumentation mean that more
sophisticated tests will be delivered to clinical diagnostics for
patient benefit. For me that’s exciting and highly motivating.
Steve Pennington was speaking to Ruairi MacKenzie, Science Writer
for Technology Networks.
9 TechnologyNetworks.com
 The Diverse Applications of Mass Spectrometry
Rapid Detection
of Designer
Drugs Presents
Challenges
for Law
Enforcement
and Forensic
Toxicology
Kimberly Scott
T
he rise of designer drugs1 is creating new challenges for
both law enforcement and forensic toxicology laboratories,
leading to new methods to identify and combat the often-
deadly substances. Designer drugs are analogs of controlled
substances that are designed to mimic the pharmacological effects
of the original drug. They range from synthetic marijuana, also
called “spice”2 to stimulants known as “bath salts”3 to fentanyl
analogs4 that are thousands of times more powerful than
pharmaceutical fentanyl and that can kill in very small doses.
George Behonick, PhD, laboratory director and chief
toxicologist at Axis Forensic Toxicology in Indianapolis, says
the most pressing focus today is on fentanyl analogs, which have
proliferated in recent years. “Some of these drugs seem to just
show up overnight,” he says. “We are now testing for about a
dozen of these analogs.”
Technology Networks 2018
According to the National Institute on Drug Abuse, there are at
least 16 fentanyl analogs, including acetyl fentanyl, carfentanyl
and cyclopropofentanyl.5 The Centers for Disease Control and
Prevention (CDC) estimates that drug overdose deaths exceeded
60,000 in 20166 and were partially driven by a fivefold increase in
overdose deaths involving synthetic opioids, including fentanyl
analogs. Carfentanyl (carfentanil) which is 5,000 times as potent
as a unit of heroin and 10,000 times as potent as morphine,7 is
one of the deadliest forms of these analogs.
Fentanyl Analogs Present Threats to Law
Enforcement
Not only do fentanyl analogs kill users at a high rate, but they
also are a significant threat to law enforcement personnel and
first responders. Minute amounts – equivalent to a few grains
of salt – of fentanyl can be lethal and visually can be mistaken
for cocaine or white powder heroin, says the Drug Enforcement
Administration (DEA), which issued a warning to law
enforcement personnel in June 2016 to exercise extreme caution
when handling possible fentanyl-containing materials.8
“One of the biggest challenges for law enforcement is the
immediate identification of substances that they find on a crime
scene,” notes Behonick. “They are trying to identify it in solid-
dose forms whereas post-mortem toxicology labs are trying to
combat the problem by identifying it through blood and urine.”
Police officers often use handheld spectrometers to scan unknown
substances for presumptive testing.9 Confirmatory testing is still
required by toxicology testing, but the presumptive tests can
be useful in determining treatment options for those who have
overdosed.
While the gold-standard technology for identifying drugs in the
lab remains mass spectrometry, the rapid rise of new designer
drugs challenges toxicology laboratories to constantly develop
tests or modify existing assays to identify these toxins.
10 TechnologyNetworks.com
 The Diverse Applications of Mass Spectrometry
“The challenge for us is in developing new methods for these
emerging drugs in biological matrices which are inclusive of the
pre-analytical phase, the analytical phase and the post-analytical
management and interpretation of data,” says Behonick.
QTOF Detects Fentanyl Analogs in Minute
Quantities
SCIEX’s X500R QTOF (quadrupole time-of-flight) is among
the current generation of mass spectrometers that can detect
fentanyl analogs in minute quantities. Introduced in 2015, the
X500R uses high-resolution MS technology to detect illicit
substances down to the picogram level, which can then be cross-
referenced through ChemSpider, a chemical structure database.
Phil Taylor, global marketing manager, food, environment
and forensics for SCIEX, sees mass spectrometry technology
moving from nominal mass instrumentation, such as the triple
quadrupole instrumentation, to more advanced platforms that
will provide a higher level of detail than previous technology.
“The pursuit in forensic toxicology is accuracy,” says Taylor.
“The demand from the judicial system is for accurate and concise
results. That’s what’s driving the market.”
Advancements in Rapid Detection of Fentanyl
Analogs
According to a study published in Forensic Chemistry in June
2017,10 advancements are also being made in using thermal
desorption direct analysis in real time mass spectrometry (TD-
DART-MS) and ion mobility spectrometry (IMS) as tools for
the rapid and sensitive (nanogram to picograms) detection of
fentanyl, 16 fentanyl analogs and five additional opioids (heroin,
U-47700, buprenorphine, methadone and naloxone).
TD-DART-MS is sensitive to picogram levels of a wide range
of illicit drugs, reports the study by NIST researchers, noting
that these instruments have potential applications in mobile
laboratories, emergency vehicles and hospitals.
“Current guidelines recommend an enzyme-linked
immunosorbent assay (ELISA) screen for fentanyl followed by
gas chromatography/mass spectrometry (GC/MS) analysis,”
write the study authors. “As NPF concentrations in blood can
be quite low, a wipe-based technique, such as TD-DART-MS
targeting solid trace contamination on the individual or their
belongings, may be a more effective approach. TD-DART-MS
may also be useful in emergency medicine, providing a rapid
identification of the specific NPF to make informed choices
about treatment.”
Technology Networks 2018
IMS instruments are commonly used in airports, where a security
officer might swab a piece of luggage or a passenger’s hands, and
then insert the swab into the instrument to check for traces of
explosive residue.
“Currently, police officers have to handle drugs to test them,” says
Edward Sisco, a research chemist at NIST and one of the lead
authors of the NIST study. “But with these technologies, they
can just swab the outside of the bag to test for fentanyl.” If the
test comes back positive, they can take extra precautions.
IMS instruments cost around $25,000 and are about the size of
a microwave oven, small enough to be transported by a mobile
hazmat unit. TD-DART-MS instruments, which are more
sensitive but larger and more expensive, could potentially be
used for screening incoming material at a forensic lab before it is
handled by evidence examiners.
NIST was the first to publish the IMS and TD-DART-MS
signatures for the 16 fentnyl analogs tested. According to NIST,
Sisco and his co-authors are speaking with IMS manufacturers
about adding the newly identified signatures to their product
software.11
“We hope this makes a real difference to the people who come
into contact with synthetic opioids,” says Sisco. “The opioid
epidemic is a huge problem. This might be one small way to try
to get a handle on it.”
References
1. The New Wave of Designer Drugs: A Review for Criminal Justice
and Forensic Professionals. Link: https://bit.ly/2JtwzWH
2. National Institute on Drug Abuse; Synthetic Cannabinoids (K2/
Spice). Link: https://bit.ly/2uhrZhG
3. National Institute on Drug Abuse; Synthetic Cathinones (“Bath
Salts”). Link: https://bit.ly/22Dcs8e
4. Drug Enforcement Agency; Fentanyl FAQs. Link: https://bit.
ly/2KSidMM
5. National Institute on Drug Abuse; Emerging Trends and Alerts.
Link: https://bit.ly/29J2oFb
6. CDC; Drug Overdose Deaths in the United States, 1999–2016.
Link: https://bit.ly/2pcubtA
7. Comparing the Lethality and Potency of Opioid Drugs. Link:
https://bit.ly/2JrDzDj
8. Drug Enforcement Agency; DEA Warning to Police and Public:
Fentanyl Exposure Kills. Link: https://bit.ly/1rh3kbt
9. Powerful Detection Technology for Powerful New Street Drugs.
Link: https://bit.ly/2iTLqtr
10. E. Sisco et al., Forensic Chemistry. 4, 108–115. (2017)
11. Fentanyl Can Sicken First Responders. Here’s a Possible Solution.
Link: https://bit.ly/2Bpo9bj
11 TechnologyNetworks.com
 The Diverse Applications of Mass Spectrometry
How Advances
in Mass
Spectrometry Are
Helping the Food
and Beverage
Industry
Dr Jenny Nelson, Assistant Adjunct Professor
of Viticulture & Enology at the University of
California, Davis, and Applications Scientist at
Agilent Technologies.
Karen Steward, PhD
M
ass spectrometry is an important tool for many industries,
not least in ensuring the safety of our food and beverages.
Hazards ranging from pesticides, to heavy metals and
toxins can be identified using the technology, even if they are
only present in minute quantities. Here Jenny Nelson tells us
how advances in mass spectrometry (MS) have improved analysis
potential and the important role it has played and will continue to
play in ensuring the safety of the beverage industry.
Q: What advances in MS have had the biggest
impact on your research?
A: Inductively coupled plasma triple quadrupole mass
spectrometry (ICP-QQQ ) has been a game changer. ICP-QQQ
provides MS/MS capability which means we can remove spectral
interferences with increased reliability and effectiveness through
more controlled chemical reactions. MS/MS improves detection
Technology Networks 2018
limits across the board, as well as providing access to isotopes
and elements that we couldn’t measure before with conventional
quadrupole ICP-MS instruments.
Q: A lot of your recent work has focused on the
beverage industry, could you tell us about some of
the projects you have been involved in?
A: Our two most recent projects working on beverages have been
with the Food and Drug Administration (FDA) studying arsenic
speciation in wine. Arsenic exists in multiple forms in foods and
beverages and not all forms have the same toxicity. The inorganic
forms of arsenic (As(III) (arsenite) and As(V) (arsenate)), are
the most toxic. Inorganic arsenic species are categorized as class
1 carcinogens. The FDA has established an action limit for
inorganic arsenic in apple juice of 10 μg/kg (ppb), however, there
are no regulations controlling the arsenic content of wine in the
US. There are limits however in the EU and Canada. Europe
(International Organisation of Vine and Wine, OIV) have set
maximum acceptable limits for total arsenic in wine of 100 µg/L
(ppb), and Canada (Vintners Quality Alliance VQA, Ontario)
have set a limit of 200 µg/L (ppb). The first project was to extend
the existing HPLC-ICP-MS method for determination of four
arsenic species in fruit juice - US FDA Elemental Analysis Manual
(EAM) 4.10 - to include wine. This work was recently published1
in the Journal of Agricultural and Food Chemistry. The second
was to develop an arsenic speciation method 10× faster than the
current FDA 4.10 regulatory method. The methodology used in
this study analyzed the inorganic arsenic species in the As(V)
form. As(III) was intentionally oxidized to As(V) with hydrogen
peroxide before analysis. By converting As(III) and analyzing
all inorganic species as As(V), this method was able to separate
from the other arsenic species monomethylarsonic acid (MMA)
and dimethylarsinic acid (DMA). The total separation time was
less than 2 minutes. This study2 utilized oxygen reaction gas in
the collision/reaction cell (CRC) of the ICP-QQQ to resolve the
spectral interferences on arsenic. Some of the interesting results to
readers would be the data on the market basket study.
12 TechnologyNetworks.com
 The Diverse Applications of Mass Spectrometry

Market
Basket Alcohol Sum of
Style Cultivar Region Vintage iAs DMA MMA
Wine (%v/v) Species
Sample
Cabernet 17.13 ± 0.83 ± 17.96 ±
S1 Red North Coast 2009 13.5 <LOD
Sauvignon 0.22 0.03 0.25
Appellation 7.49 ± 0.30 ± 0.77 ±
S2 Red Pinot Noir 2004 13.8 8.56 ± 0.52
Central Coast 0.15 0.062 0.32
Santa
14.63 ± 0.80 ± 15.43 ±
S3 White Chardonnay Barbara 2013 13.5 <LOD
0.40 0.08 0.48
County
Napa and 25.03 ± 0.69 ± 0.47 ±
S4 Rose Zinfandel 2013 10.5 26.19 ± 1.27
Sonoma 0.89 0.26 0.12
23.45 ± 0.32 ±
S5 White Chardonnay Central Coast 2013 13.5 <LOD 23.77 ± 1.17
1.12 0.05

The table above shows quantitative results (μg/kg) for inorganic and organic arsenic species in five commercially available wines measured by
LC-ICP-QQQ. In all cases, most of the arsenic was present as the inorganic forms (iAs). However, the arsenic levels were all much lower than the
maximum allowable level of total arsenic in wine defined in Canada and Europe.

Q: What are the biggest challenges you face when
applying mass spectrometry to food and beverage
analysis?
A: Historically, the perception of ICP-MS has been that it
is complicated and difficult to use, presenting its own set of
challenges in the application of mass spectrometry to food
and beverage analysis. However, modern ICP-MS instruments
are much more user-friendly, with built-in auto-optimization
and diagnostics capabilities. ICP-MS now provides analytical
possibilities that extend the lab’s capability beyond what’s offered
by the more traditional techniques we previously used.
In fact, ICP-MS greatly simplifies the analysis of foodstuffs,
world of ICP-MS, enabling us to now develop methods around
elements/metals that used to be challenging. One good example
of this was a collaboration where we worked with Brian Jackson
at Dartmouth University. The aim of this study3 was to accurately
measure arsenic and selenium in foods using Triple Quadrupole
ICP-QQQ to remove doubly charged rare earth element (REE)
interferences. REE levels in food samples are low, but crops
grown in REE-enriched soils have the potential to have higher
concentrations of these elements, leading to false positives for
arsenic and selenium. It is known that arsenic and selenium can be
difficult to quantify accurately by conventional single quadrupole
ICP-MS, especially at low levels. This is because isotopes can
suffer from multiple spectral interferences, from doubly charged
ions of the REE (REE ), as well as polyatomic ions formed from
++

compared to the traditional analytical approaches. This would the sample matrix and plasma. A quadrupole mass spectrometer
typically have involved AAS or ICP-OES for the major and separates ions based on mass-to-charge ratio (m/z), and so the
nutrient elements, graphite furnace AAS for the toxic and trace REE ions appear at half their true mass, overlapping the singly
++

elements, and a dedicated analyzer for mercury. All these analytes
can now be measured – at much lower detection limits – in a
single, fast, practically interference free ICP-MS measurement.
So, the potential sample throughput with ICP-MS is much greater
than before, and the cost per analysis is far lower. Plus, the method
simplification in terms of sample preparation, calibration strategy,
QC overheads, reporting, waste disposal etc., gives concrete
benefits especially in commercial food labs.
Q: How do you think developments in mass
spectrometry are likely to change the food and
beverage industry in the future?
A: The removal of spectral interference provided by the ICP-
QQQ system through MS/MS, has really changed things in the
charged analyte ions of arsenic and selenium. This table shows a
comparison of single quadrupole (SQ) and triple quadrupole ICP-
MS results for arsenic and selenium in food reference materials.
Significant errors in the (uncorrected) single quadrupole results
meant that correction equations had to be applied to improve the
accuracy. Triple quadrupole ICP-MS provides better accuracy
without requiring any correction equations, illustrating an
important advantage of ICP-QQQ data vs. traditional SQ ICP-
MS for this type of interference.
Additionally, analysis of nanoparticles in food is an emerging
application, the development of which has greatly benefited from
the advances in single particle inductively coupled plasma-mass
spectrometry (sp-ICP-MS). As well as advances in the hardware
in the ICP-MS, there have also been new software capabilities

Technology Networks 2018 13 TechnologyNetworks.com

 The Diverse Applications of Mass Spectrometry

ICP-QQQO2/
ICP-MSHe mode ICP-MSH2 mode
H2 mass-shift
SRM Certified Uncorrected Corrected Uncorrected Corrected Uncorrected
Arsenic (mg/kg)
NIST 1547 0.060+0.018 0.170+0.016 0.068+0.003* 0.113+0.004 0.079+0.004* 0.065+0.002*
NIST 1515 0.038+0.007 0.250+0.016 0.026+0.021* 0.126+0.005 0.047+0.004* 0.032+0.002*

Selenium (mg/kg)
NIST 1547 0.120+0.009 0.394+0.04 0.113+0.02* 0.119+0.009* 0.119+0.009* 0.127+0.006*
NIST 1515 0.050+0.009 0.808+0.04 0.013+0.04* 0.050+0.003* 0.050+0.003* 0.047+0.006*
*95% confidence interval overlaps with the certified range

that have had to be developed to support this new application. References

I believe that advances in ICP-MS instrument sensitivity and 1. C. Tanabe et al., J Agric Food Chem. 65, 4193-4199 (2017)
sp-ICP-MS software will lead to this type of measurement soon 2. P. J. Gray et al., J. Anal. At. Spectrom. 32, 1031-1034 (2017)
becoming mainstream; regulators are already looking at how these 3. B. P. Jackson et al., J. Anal. At. Spectrom. 30, 1179-1183 (2014)
novel nanomaterials should be monitored and controlled. Since
this is a new hot topic, I foresee many advances in technology
from the instrument vendors in the future as they gain a better
understanding of the requirements of both their customers and
the industry.

Dr Jenny Nelson was speaking to Dr Karen Steward, Science Writer for

Technology Networks.

Jenny Nelson has been an Adjunct Professor in the Department of

Viticulture and Enology at University of California, Davis since 2013 and
is an Atomic Spectroscopy Research Scientist for the Applied Markets team
at Agilent Technologies. Her research focus is in trace elemental analysis for
the food and beverage industry. She has extensive experience in operating
and method development for Inductively Coupled Plasma Mass Spectroscopy
(ICP-MS) and Inductively Coupled Plasma Optical Emission Spectroscopy
(ICP-OES), and speciation analysis for many sample matrices using GC-
ICPMS and LC-ICPMS.

Technology Networks 2018 14 TechnologyNetworks.com

 The Diverse Applications of Mass Spectrometry

Measuring
How the Brain
Metabolizes in
Real-time
Adam Tozer, PhD

P
reviously, if you wanted to analyze brain metabolites
over time in a mouse study, you would need to extract
brains from different mice at different time points. An
‘upside’ to this method is that metabolite changes could be
measured from whole areas of the brain. However, the data only
provided a snapshot of the metabolic changes at distinct time
points. Previously, it was not possible to view metabolic changes
happening in real-time. Ground-breaking work included
the development of checkpoint blockade therapy,5 targeting
cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4)6
and programmed death receptor-1 (PD-1)7 with monoclonal
Figure 1: Differences between conventional metabolome analysis for dissected brain
antibodies in tumors. Many patients treated with these new
samples and the newly developed in vivo real-time monitoring system. Conventional
therapies had tumor remissions which has raised the prospect metabolome analysis for dissected brain samples is widely accepted as a general
that some of these cancers may be manageable as chronic protocol, though there is a critical issue: the effect of death on metabolome cannot
conditions. be avoided. However, the newly developed in vivo real-time monitoring system
can observe metabolic dynamics without the effect of death and there are high
An innovative approach developed by Assistant Professor Kei expectations for it to reveal hidden mechanisms. The system will also provide a novel
Zaitsu and Lecturer Yumi Hayashi at Nagoya University, Japan, visualization technique for homeostasis, which can be applied in healthcare risk
enables the real-time monitoring of brain cell metabolism in management. Credit: Kei Zaitsu
anaesthetized animals.
By combining an acupuncture needle as an ionization device
Needle-sharp Sampling and Identification
 with sensitive mass spectrometry machinery, the team
successfully monitored eight cerebrum metabolites related to
The team previously showed that the new system could be used central energy metabolism at 20 second intervals for three hours,
to monitor metabolomic changes in the liver1 of anaesthetized in anaesthetized mice. The findings are published in the journal
rats in real-time, so next wanted to try the system on the brain. Analytical Chemistry.2

Technology Networks 2018 15 TechnologyNetworks.com

 The Diverse Applications of Mass Spectrometry
Figure 2: Schematic image of a freely-movable stage. The newly developed stage is composed of
an x-y-z freely movable stage, rubber heater, and fixing arm unit. The stage enables control of
the sampling point with μm-order spatial resolution. Credit: Kei Zaitsu.
Co-lead author, Kei Zaitsu explains how the system works:
“PESI/MS/MS stands for probe electrospray ionization (PESI)
with triple quadrupole tandem mass spectrometry (MS/MS)” 
 sensitive proteins into brain cells, is a powerful way to probe
“The thin needle (tip diameter, 700nm) moves into and out of
the tissue. The tip performs the sampling, whilst the top of the
needle is the ionization unit.”
“The ionized brain metabolites then get fired into the mass
spectrometer for quantification.”
 pairing. Kei admits this is something he would like to do:
The need for the tandem quadrupole tandem mass spectrometry
(MS/MS) in this set up is explained by Yumi Hayashi:

“While the thin probe needle in PESI enables direct sampling
with subcellular-level invasiveness, and combinational use
of PESI and single mass spectrometry (PESI/MS) has been
applied to direct analysis of target compounds, PESI/MS lacks
selectivity because PESI does not possess chromatographic
separation property,”

 near future.”
Yumi adds: “Therefore, combinational use with a specific
detector such as MS/MS must be explored for reliable
identification of metabolites.”
Real-time Information
The technique affords unprecedented insight into the
metabolism of a small area of brain tissue in real time. Whilst Kei
disclosed that there is no way to know how many cells are being
sampled from at any one time, at 700nm the tip is small enough
to investigate specific regions of brain nuclei, or cortical layers,
Technology Networks 2018
for example. This is a marked improvement in the specificity of
brain metabolomics.
The system is not only useful for metabolic measurements, as
Kei explains: “We know the dynamics of metabolites in a living
mouse’s brain. There is also a high possibility to monitor the
dynamics of peptides and neuropeptides in real-time in the
future.”


The system provides unprecedented access to neuroscientists
and chemists alike. Understanding the flux in metabolites and
neurochemical messengers in disease models could add much-
needed insight to the development pathologies and provide new
opportunities for therapeutic interventions.
Real-time Metabolomics Could be Combined with
Other Techniques
Right now, neuroscientists can use electrophysiology to explore
the electrical activity of brain cells. Combining electrophysiology
with optogenetic techniques to enable activation and silencing
of brain cells with light, by genetic incorporation of light-
brain function and behavior.3 Scientists also use chemogenetic
techniques to manipulate brain cell activity with designer
receptors and ligands.4
Combining real-time metabolomic measurements with real-
time manipulation of brain cell activity would prove a powerful
“Yes, we would strongly like to combine this technique with
optogenetics.”


Whilst the advance of getting metabolomic data in tandem with
brain cell activity data is desirable, neuroscientists would want
to achieve this in non-anaesthetized mice and possibly combine
the data with a behavioral test. Something Kei also recognizes:
“The mouse needs to be anesthetized at this time, but we will
modify the system to be applied to freely moving mice in the
Advancing Drug Discovery and Research
In the paper, the scientists tested the effect of cannabinoid type-1
receptor agonist on brain metabolomics. This proof-of-concept
study showed how the PESI/MS/MS system could be used
in drug discovery, possibly as part of toxicological profiling in
animal models. The technique would enable detailed changes in
brain metabolism caused by drugs to be measured in real-time.
The technique can be applied to a host of different research
situations and PESI/MS/MS could revolutionize our
16 TechnologyNetworks.com
 The Diverse Applications of Mass Spectrometry

understanding of the function of many tissues in health and

disease. As Kei explains: “The demonstrated capabilities of the
new PESI/MS/MS system make it a promising tool for the
analysis of neurodegenerative diseases, such as Alzheimer’s
disease. We can also use it to analyze other tissues such as liver
and kidney, suggesting a wide-range applicability of the present
system,”

As a pioneer of PESI/MS/MS, Kei is right to be excited about

this technique and hopes it will spawn a new field of ‘real-time
metabolomics’.

References:

1. K. Zaitsu et al., Anal. Chem. 88, 3556–3561. (2016)
2. K. Zaitsu et al., Anal Chem. 90, 4695–4701. (2018)

3. N. Yamawaki et al., Cold Spring Harb Protoc. 10, (2016)

4. K. Berglund et al., Methods Mol Biol. 1408, 207–225. (2016)

Technology Networks 2018 17 TechnologyNetworks.com

 The Diverse Applications of Mass Spectrometry
Mass
Spectrometry’s
Important Role
in Identifying
Explosives in the
Environment
Professor Stanislaw Popiel, Associate
Professor of Chemistry at the Military
University of Technology, Warsaw, Poland.
Karen Steward, PhD
E
xplosives, especially when not contained or disposed of
correctly, pose a significant hazard to humans, wildlife
and the ecosystem. Having the tools to be able to identify
their presence and state are therefore key to combatting the issue.
We spoke to Stanislaw Popiel about the game-changing role
that mass spectrometry is playing in the process and some of the
challenges that still lie ahead.
Q: Could you tell us a bit about the role and
importance of mass spectrometry in explosive
screening?
A: Explosives screening is very important in a number of
situations including terrorism attacks, environmental monitoring
(training grounds, factories, ground waters etc.) and dumped
munitions. Mass spectrometry gives us a powerful tool for
identification and quantification of explosives, vital in forensic
analysis.
Technology Networks 2018
As well as the explosives themselves, their degradation products
can also be detected, which may indicate their use. Tandem
mass spectrometry allows us to reduce background signal
from the sample matrix and reduce the limit of detection and
quantification.
Q: How do the analyses made possible through
the application of mass spectrometry compare to
previously available methods of detection?
A: Mass spectrometry in combination with gas chromatography
and especially liquid chromatography can be used in a very
wide research spectrum related to explosives. In comparison to
other techniques, mass spectrometry is advantageous because
with one detector we can do both qualitative and quantitative
analysis of explosives and their degradation products. Also, mass
spectrometry, especially high-resolution mass spectrometry,
enables us to monitor the degradation and metabolization path of
explosives, and identify unknown compounds.
We believe that our work on the development and validation
of methods for analysis of all explosives and their degradation
products will allow for their practical use in environmental
monitoring and forensic science. In addition, the possibility
of substance identification gives us the opportunity for more
complex analyses of environmental or forensic samples.
Q: What are the biggest challenges that are still to
be overcome and how might they be addressed?
A: The biggest challenge is to develop and validate sensitive
methods for the detection of explosives and their degradation
products in sea water and sediment samples, as large amounts of
conventional and chemical munitions are dumped in the oceans
and seas. Both chemical warfare agents and explosives undergo
very complex transformations under the influence of the marine
environment. As a result, after a long time, a lot of substances
significantly differ from the starting substances which were
dumped in the marine environment. In addition, substances
18 TechnologyNetworks.com
 The Diverse Applications of Mass Spectrometry

appearing as a result of chemical or biochemical transformation References

are on the one hand in very different concentrations, and on
the other hand, often have similar properties and therefore it is
difficult to distinguish, identify and quantify them. Sea water and
sediments are very complex matrices and extraction of analytes
with one solvent yielding good recovery is difficult to achieve.
1. P. Lebel et al., J. Chromatogr. A. 1343 134–151. (2014)
2. J. Kim et al., Anal. Bioanal. Chem. 408, 251–263. (2016)
3. F. Shi et al., J. Chromatogr. A. 1344, 91–98. (2014)

A related significant aspect is to determine the influence of

explosives and their degradation products on the marine
environment. In both cases, mass spectrometry is a powerful
tool and importantly does not limit identification to known
metabolites and degradation products. This information is
crucial for determining the toxicity of explosives and allows us to
trace the path of their metabolism.

Q: Can you tell us about an occasion where this

technology has been successfully applied to
combat crime?

A: Our laboratory have applied mass spectrometry for the

identification and quantification of explosives and their
degradations products in sediment samples and actual explosives
fished out of the Baltic Sea. These have included trinitrotoluene
(TNT) and (cyclotrimethylenetrinitramine) RDX.

Mass spectrometry has crime fighting applications beyond

explosives including the analysis of drugs1, 2, herbal medicines3
and dietary supplements3. These applications are very important
in forensic analysis because their results are used as evidence
in criminal proceedings. Mass spectrometry gives us new
possibilities to search for trace substances that can be used as
evidence.

Professor Stanislaw Popiel was speaking to Dr Karen Steward, Science

Writer for Technology Networks.

Dr. Stanislaw Popiel is an Associate

Professor of Chemistry at the Military
University of Technology in Warsaw,
Poland. Prof. Popiel has over 30 years
of experience in analytical chemistry,
especially in analysis of dangerous
substances, such as chemical warfare
agents, explosives and their degradation
products. His main task is the
development and validation of analytical methods based on both gas and
liquid chromatography especially in combination with mass and tandem
mass spectrometry. In his lab, Prof. Popiel has a number of analytical
instrumentations with several Master’s & PhD students. He holds
numerous publications in his field and notable awards.

Technology Networks 2018 19 TechnologyNetworks.com

 The Diverse Applications of Mass Spectrometry
Mass
Spectrometry in
Biopharmaceutical
Discovery
Rachel Brazil
B
iotherapeutics now makes up 25% of new drugs
approvals. Their efficacy and safety are highly dependent
on their structures, which are complex, heterogeneous
and subject to modification. Analysis of biological molecules
therefore needs a different approach to small molecule
pharmaceuticals. Mass spectrometry has become the go-to
technique to supply answers to a range of analytical questions,
offering sensitivity, selectivity and specificity. Over the past
decade advances in mass spectrometry are allowing more
information on higher order structure and new imaging
techniques are even providing insights into how biological drugs
behave in vivo.
Mass spectrometry (MS) can detect, identify and quantify
molecules separated by their mass to charge ratio. But biological
molecules pose some unique problems. “They are often hundreds
of times the size of conventional small molecule drugs,” says
Todd Stawicki, biopharma applications scientist at SCIEX.
This tends to affect the sensitivity you can achieve with mass
spectrometry. Plus, “biosynthesis is highly heterogeneous,” adds
Stawicki, “it often creates lots of small variations. Then the
analytical challenge is to characterise those small variations and
determine if they are clinically relevant.”
Technology Networks 2018
Typically, a mass spectrometer ionizes the sample using
electrospray ionization (ESI) where a high voltage is applied
to a liquid to create an aerosol. This ‘soft ionization’ technique
creates little initial fragmentation – often an issue with large
molecules. Ions are separated by acceleration in an electric or
magnetic field and then detected by an electron multiplier: the
deflection experienced is a function of the mass-to-charge ratio.
The mass-to-charge ratio of the ions is determined by one or
several different types of mass analyzers. To gain more structural
information tandem mass spectrometry (ESI-MS/MS) is used,
where discrete ions can be isolated, fragmented and their mass-
to-charge ratio determined.
Mass Spectrometry and Protein Structure
The information that can be obtained from a protein is immense,
including its primary amino acid sequence, post-translational
modifications and even higher-order structure. The most basic
information – its amino acid sequence – is routinely found by
trypsin digestion to create peptide fragments that produce a
fingerprint mass spectrum. Structural information on the various
polypeptides within a protein can also be found using methods
such as breaking disulphide links using a reducing agent. “With
an antibody you can simply reduce them, so rather than one
single protein you are going to get four smaller subunits. It can
allow you a finer resolution and detail while still maintaining
a lot of its intact structure,” says Kelli Jonakin, Senior Global
Marketing Manager for Pharma and BioPharma at SCIEX.
“There are also a wide range of different post-translational
modifications that can be directly observed by mass
spectrometry,” says Jonakin. These are the enzymatic
modifications that occur to proteins following biosynthesis and
can have an impact on the efficacy and safety of a drug. An
important one is glycosylation – the covalent addition of sugar
moieties to specific amino acids. Approximately half of all
proteins expressed in a cell undergo this modification. Glycans
20 TechnologyNetworks.com
 The Diverse Applications of Mass Spectrometry
can be enzymatically separated from the protein before analysis
to produce a glycosylation fingerprint. By engineering a protein’s
surface glycosylation pattern, drug developers hope to enhance
therapeutic performance.
Mass spectrometry also has a crucial role in detecting
contaminants from the bioengineering process says Jonakin:
“‘When the engineered cells are producing these biotherapeutics
they also produce lots of other proteins (host cell proteins,
HCPs) and some of these can have undesirable properties.” Mass
spectroscopy is able to characterize and quantify these impurities.
“In development, if there is a host-cell protein that is particularly
immunogenic and represents a high safety risk, scientists could
develop an LC-MS assay to quantitatively monitor a unique
signature peptide for that host-cell protein, and then monitor for
this in the purification and release batch for the biotherapeutics,”
explains Jonakin.
Mass Spectrometry and Drug Efficacy
In biopharma mass spectrometry play a role in measuring
drug efficacy often by analyzing downstream effects. “We can
employ it for looking at the biological consequence of the drug-
target binding,” says Stawicki, “for example, we can use mass
spectrometry to look at phosphorylation of a messenger protein
caused by the binding event.” But, it is also now possible to probe
a biotherapeutic’s interactions with its target by performing a
native mode analysis.
According to chemist, Igor Kaltashov, from the University
of Massachusetts Amherst, the technique is not yet widely
established, but he says: “industry is becoming more interested
in this aspect of mass spectrometry. I would say in 2–3 years it
will become commonplace.” Native mass spectrometry requires
protein assemblies to be extracted from solution into the gas
phase using ESI. “You have to do it gently, so you don’t break up
these complexes.” To assist the move from solution into the gas
phase Kaltashov pioneered a technique that uses size exclusion
chromatography. This first allows the molecular complexes in
solution to be separated from smaller molecules whilst preserving
biological activity. “You can inject your complex as it’s formed in a
phosphate buffer using a solvent system that isn’t compatible with
mass spectrometry and use size exclusion chromatography as an
interface,” explains Kaltashov.
Another technique to study conformation and binding is
hydrogen-deuterium exchange (HDX) MS. If heavy water
is introduced in solution, hydrogen atoms will exchange with
deuterium. The rate of exchange is characteristic of the
degree to which the hydrogen is protected and that provides
conformational information. “The binding epitope (i.e. the
interface residues) will be shielded from your labeling agent and
Technology Networks 2018
so once the labeling is completed you can interrogate them and
by using standard approaches, determine which residues have not
been labeled. They are assigned as the residues that are involved
in the formation of those binding epitopes,” explains Kaltashov.
The Kit
Many types of studies require highly sensitive instruments
and Jonakin says the gold standard for protein quantification is
the triple quadrupole mass spectrometer (triple quad, TQMS,
QqQ). This is a tandem mass spectrometer consisting of two
quadrupole mass analyzers in series. Quadrupole refers to it being
constructed from four parallel cylindrical rods which control the
voltage and allow only ions of a given mass-to-charge ratio to
reach the detector. In between the two mass analyser quadrupoles
is a third quadrupole which acts as a cell for collision-induced
dissociation. “Triple quads are extraordinarily sensitive and there
are many places within the discovery and development pipeline
where researchers need extreme sensitivity,” says Jonakin, ‘this
can be critical for pre-clinical and clinical studies. But, they do
not provide deep MS information, that is what quadrupole time-
of-flight MS gives you.’’
Quadrupole time-of-flight MS (Q-Tof MS) is another important
high-resolution mass spectrometry method. In a time-of-flight
mass analyzer, ions are accelerated in an electric field. The pulsed
ions travel in a high vacuum, field-free region and then impact an
electron multiplier. Ions travel as a function of their size – small
ions travel faster than large ions. “This allows us to detect and
characterize proteins and peptides with extremely high accuracy
and resolution. That is critically important for confidently
distinguishing very small, yet important, differences in very
big biotherapeutics,” says Stawicki. These state-of-the-art mass
spectrometers can offer a level of detailed information that, for
example, enables accurate glycosylation profiling.
For studying biological drugs, Jonakin says Q-Tof MS systems
provide a speed advantage: “one of the greater needs for
biopharmaceuticals has been speed, pretty much at all levels,
because speed equates to depth of coverage and because
biologics are so complex you just have so much more going on.
The ability to do a lot of experiments in a short time frame
allows you to get very deep and very rich mass spectrometry
information.”
New data acquisition strategies are also allowing much more
comprehensive fragment ion analysis in biological samples using
high-resolution Q-Tof mass spectrometry. This can give you
detailed information on every detectable analyte in the sample
in a single run. For highly complex biotherapeutics this is a great
advantage.
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 The Diverse Applications of Mass Spectrometry
Mass Spectrometry and Imaging
Another innovation that has proved useful in biopharmaceutical
development is imaging mass spectroscopy. One of the earliest
methods used for biological imaging was MALDI (Matrix-
assisted laser desorption ionization) imaging, first developed in
the 1990s by Richard Caprioli at Vanderbilt University. Twenty
years later its use is becoming widespread for imaging tissue
as part of pharmacokinetic studies. “The thing about MALDI
compared to some other imaging MS methods is that it is able
to see large biomolecules such as peptide and proteins,” says
Adam McMahon, Senior Lecturer in Analytical Chemistry at the
University of Manchester.
The method works by covering a thin tissue section with a
crystallised matrix material and progressively scanning the
surface with a laser. This evaporates the matrix which carries
the underlying biomolecules with it. “You generally see intact
molecular ions and not their fragments,” says McMahon. “Image
files can contain tens of gigabytes of data in which you have
peaks at very many masses, each of which will give you an image.
So, if you have a drug molecule, you can potentially map its
distribution across the tissue sample and can see whether your
drug has penetrated into a particular tissue region, which might
be important in a tumor, for example.”
Such imaging has been used to explain some unusual off-target
drug side effects. For example, MALDI was used to identify the
drug metabolite that caused seizures in patients taking an HIV
non-nucleoside reverse transcriptase inhibitor. “Drug delivery
into the brain can also be examined, where the blood brain
barrier can exclude certain molecules,” adds McMahon. Others
have been using the technique to determine tumor margins,
comparing MALDI MS images to histology data.
Technology Networks 2018
Another imaging method, desorption electrospray ionization
(DESI), instead of using a laser, uses a stream of fast moving
charged solvent droplets in the order of 1 - 5 microns in size,
aimed at the sample surface. The droplets extract molecules from
the surfaces into the gas phase for analysis. McMahon says there
had been scepticism about the spatial resolution the technique
could provide, but it can now be as low as 40–50 microns; “You
do not see the large proteins extracted in these droplets, you see
lipids mainly. But lipids can tell you an awful lot about what is
going on in your tissue. You may be comparing a tissue that has
and has not been treated with a drug and what you see is the
tissue response in terms of the change in the lipid profile.”
Mass spectrometry is a crucial tool in the discovery and
development of biological drugs but to many is often still seen
as a technique for specialists, with results needing complicated
interpretation. Developments and subsequent advancements in
mass spectrometry mean that the technique is now a lot more
accessible to researchers that are less experienced in using the
technique.
The power of mass spectrometry is now something that all
researchers in biotherapeutics can harness themselves.
References
1. V. Dotz et al., Trends Analyt Chem. 73, 1–9. (2015)
2. K. Muneeruddin et al., Anal. Chem. 86, 10692–10699. (2014)
3. C. E. Bobst et al., Anal. Chem. 80, 7473–7481. (2008)
4. S. Castellino et al., Chem. Res. Toxicol. 26, 241-251. (2013)
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 The Diverse Applications of Mass Spectrometry
Helping Analytical
Chemistry
Embrace Big Data
Ruairi MacKenzie
M
any recent advances in research have aimed to
maximize the amount of data we can produce. With
the cost of data handling storage plummeting, why
wouldn’t you? But as anyone who has spent hours pipetting
with an uncalibrated pipette or watched all 29 series of the
Simpsons will tell you, quality is more important than quantity.
This realization has hit home in many companies that are
now buried under a pile of poorly-stored data that can’t be
synchronized to other data silos and is occupying many
terabytes of storage. In analytical chemistry, that data has more
complexity and value than everyday spreadsheets, and tools
matching that complexity will be needed to get data back into
shape.
“If you don’t have correct data then it’s pretty much unusable
by anybody downstream, including yourself, for anything
that you originally intended it for,” says Andrew Anderson,
Vice President of Innovation and Informatics Strategy at
Toronto-based analytical software supplier ACD/Labs.
Anderson suggests that this need for correctness is now being
recognized at the beginning of the data life cycle – and the end:
“Organizations like the Food and Drug Administration require
pharmaceutical companies and drug manufacturers to have
safe, efficacious and quality drugs and the data that they supply
for characterizing those drugs has to meet guidelines according
to data integrity. There’s both the pragmatic impetus right from
the get-go and at the end. What is the expectation if you’re
going to bring a product to market that is supposed to benefit
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people? If it’s not what it’s supposed to be, there could be really
serious consequences.”
Anderson’s view is that data integrity is important at all stages
of the research pipeline, from design to drug. This perspective
has become vital as advances in technology enable data to be
recorded from more sources in larger volumes: “One of the
trends in industrial innovation is utilizing what we would call
the secondary or tertiary value that you would get from data.
Historically, if you look at how analytical data is leveraged
within industry, it’s question and answer, input and output.
What people have recognized is that by having data you can
infer trends, you can apply and use data for training sets, or
things like predictive analytics, machine learning and the like.
If I’m using analytical data to release a substance to be used in
a pharmacy setting or in a commercial setting, that released
data is used to give a green light to say, yes, you can release the
batch for its intended use. If you store that data right on every
batch that’s ever been released, you can look at trends, and infer
operational optimization decision making – do I see any trends
in how quality at one site differs from another, for example?”
With these potential benefits available, it’s surprising that
analytical chemistry has been slower than other fields to
embrace big data techniques, with available datasets and
algorithms often not up to the task of analyzing complex
chemical data. Andrew’s colleague, and ACD/Labs’ Director
of Strategic Partnerships, Graham McGibbon, says that the
complexity and volume of data are the biggest obstacles to
simply adopting automation techniques: “You have optical
spectra across ranges of wavelengths, you have experiments
performed not just for the certain sampling frequency
but across all frequencies. It takes time to run them – a
chromatography run could take half an hour. If you’re acquiring
data for that entire half hour and you have a mass spectrometer
attached, there could be thousands or millions of data points.
Furthermore, you have multiple dimensions of information
where you can probe how atoms are attached to each other.
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 The Diverse Applications of Mass Spectrometry

People want to know which peaks represent which atoms recognize - the nature of where data is assembled and what
or features, and that complexity is really a key thing about trade-offs there are in terms of getting complete and accurate
chemistry data. I think it’s much more complicated or complex data and making it useful.”
than for some other data that people would choose to store in
other fields.”
Andrew Anderson,
Andrew notes that labs or companies conducting large-scale Vice President of Innovation and Informatics
chemical analyses could end up with a mind-boggling amount Strategy, ACD/Labs.
of data: “If we want to do big data analysis, we’re generating
a terabyte of data a day, and you’re going to get to a petabyte
fairly fast over time. Being able to do the types of analyses
we’d like to do is hard if you’re not reducing the data volume
somehow.”
Graham McGibbon,
Such a deluge of data certainly sounds like a good reason to Director of Strategic Partnerships, ACD/Labs
avoid altering with a company-wide data system, but Andrew
firmly believes that even if adopting big data techniques isn’t an
easy road to walk, the alternative is far worse: “I’m personally
familiar with a situation in food and beverage companies
dealing with pesticides that had to respond to a pesticide
becoming regulated. They spent 18 months doing the hazard
assessment on their commercial products and raw material
supply chains. If you have the big data system it’s a query, a
simple query as opposed to what they had to do because the
big data systems aren’t in play. They had to gather samples, re-
analyze and go from there. If you’re going to consider adopting
big data techniques in analytical chemistry, consider the value
proposition—that’s how they justified a data center investment.
If you build this and you house it, and you architect it the right
way, it will pay off, you can avoid those 18 months’ worth of
cost to solve a problem.”

Whilst the need to advance big data techniques might seem

clear, the way in which companies choose to adopt those
techniques is less so. Who exactly has to promote more
data-centric strategies within a company? “It’s not like any
individual department gets saddled with an innovation strategy
like this, there has to be all stakeholders at the table,” says
Andrew. “You must have a concerted plan to migrate from the
current strategic set of capabilities to a new set of capabilities.
So, I wouldn’t put any single department under the gun, so to
speak, to have a responsibility to build something like this, it
has to be an inter-department function.”

Modern informatics solutions clearly have the potential to

improve how entire industries treat their data and bring
outdated practices to an end. What is also clear is that
implementing these solutions requires an intensive, but
worthwhile effort. Andrew sums up the task ahead for
companies who want to improve how they handle and analyze
data: “If somebody can mine data, then I think that’s great
but it’s an uncertain additional value compared to what they
were doing in the first place. I think that’s really important to

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 The Diverse Applications of Mass Spectrometry

Q: How did you come to work in this area of

Developing research?

A: Once I had completed my undergraduate degree I went

FTICR Mass on to do my PhD, although I originally planned to focus on

computational modelling, once I got to university that didn’t

Spectrometry
really appeal to me. And so I wandered around attempting to find
some other areas of interest, and eventually stumbled into the lab
of Fred McLafferty (the American chemist known for his work in

Instruments
mass spectrometry) who was doing very similar research to what
we are still doing now –although in a much more preliminary
state at that point – I got to know about mass spectrometry, really

with Unique
fell in love with it and stuck with it all these years. It initially
piqued my interest in the early ‘90s so roughly 25 years ago.

Q: Could you touch on your work within the area

Capabilities of cancer research?

A: I wouldn’t consider myself an expert in cancer research but

Peter B. O’Connor, Professor of Analytical my colleague, Peter Sadler, a co-author on one of our latest
Chemistry, Warwick Centre for Analytical papers works on anti-cancer metallodrugs. The most famous of
Science (WCAS), Department of Chemistry, them is cisplatin which is probably the most prescribed anti-
University of Warwick, UK cancer drug of all.

Laura Elizabeth Mason Cisplatin acts as a sort of template for most of what Peter works
on, except that he changes the metal and the ligands on the
drug – this, of course, changes the chemistry of the molecule

P
eter and his team work collaboratively with other
research groups to demonstrate the effectiveness of higher
specification fourier-transform ion cyclotron resonance
(FTICR) mass spectrometry in specific applications. Here he
discusses the diverse range of projects he is currently working on.
He touches on the adaptability of mass spectrometry, enabling
researchers to study cancer, neurological disease, petroleum, and
biopharmaceuticals.
completely. The goal is to find anti-cancer drugs which are more
efficacious, that have enhanced targetability, or, in this case,
the team wanted to find an anti-cancer drug that was photo-
activatable so that they could dose a person with an inert drug
at fairly high concentrations and then just activate the molecule
with blue light on specific target sites where the cancer is
specifically located.

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 The Diverse Applications of Mass Spectrometry
This strategy would mean that it would be possible to limit
the damage to healthy cells through controlled positioning
of the light, which is an interesting concept that Peter has
brought forward. My personal interest was directed at the
photo-activatable anti-cancer drugs, in figuring out what these
drugs were doing when they became activated. What happens
when you put this type of drug into a biological system, how
do they actually kill cells, and what are they actually doing? It
is a really interesting problem because it’s a mass spectrometry
problem. It is also a proteomics problem because most of the
drug’s interactions are with proteins in the cells. We started
out with cisplatin and found ways to detect the modification
sites of proteins in cisplatin, now we’ve extended this to
quite a few other metallodrugs. We then performed liquid
chromatography combined with mass spectrometry and tandem
mass spectrometry – so fragmentation mass spectrometry. We
used this approach to try to identify the proteins and the sites
of modification. If you do this with a conventional proteomics
method, you’ll be able to find something but everything that’s
modified with a drug has a very strange isotope pattern because
metals have very strange isotope patterns and there’s a metal at
the middle of the drug. So we had to find a way to automatically
highlight all the peaks that had the metals on them. We were
able to build a script that could examine the isotope patterns
and identify only the peaks that would have the metals on them
and we could then focus on those and perform more extensive
mass spectrometry. It is a proteomics challenge, so you have a
terrible mish mash of proteins and peptides in there, as usual.
These are also human cell lines so they’re quite complicated –
they’re not as simple as a yeast cell for example. Then we have
the metals which make such a broad isotope distribution that it’s
rather challenging to find them and that tends to mess up a lot
of the proteomics methodologies that are pretty well established
now. So, while we had to develop the tools to do that, the high
resolution of FTICR MS was very helpful, although probably
not strictly necessary in all cases. Inevitably in biological systems
you end up with overlapping isotope distributions and then that
very high resolution becomes critical. So some of this could
have been done with other proteomic set ups, but the FTICR is
clearly going to be the best approach.
Q: What are the benefits of FTICR mass
spectrometry and what applications is it
particularly suited for?
A: The biggest advantage of FTICR by far, is that it has much
higher resolution than any other type of mass spectrometer. It is
about 10 times better than the nearest competitor which is the
orbitrap. High resolution is very useful, particularly whenever
you have particularly complex samples. One of the biggest
areas that researchers work on in this area, although I don’t
work on this myself, is petroleum. Petroleum is a horrendously
complex mixture, hundreds of thousands of components, all
Technology Networks 2018
low molecular weight and so they tend to give you just huge
signal and you might have hundreds of individual peaks at
every one dalton spacing in the spectrum, so it gets really
complicated. Another area that we’re working on quite a bit is
synthetic polymers, particularly the block copolymers where
combinatorial confusion in the spectrum is common because
you have distributions of polymers that you tag together in a
block copolymer and so they get rather heterogeneous. A more
advanced mass spectrometer is necessary to make sense of the
spectra.
Both petroleum and polymers can be either not very amenable
to chromatographic separation or they can be too complicated
for chromatographic separation so you need to be able to handle
many different components at the same time in the spectrum, so
you absolutely need the resolution. Proteomics is very similar in
that sense. In this case, like in many cases, we start by digesting
the proteins in the cells down to peptides and work from there.
It’s what we call a bottom-up approach to proteomics but that
means that you’re turning maybe tens of thousands of proteins
into millions of peptides or at least hundreds of thousands of
peptides. Then you have to separate out that mixture and process
it. Now peptides are very amenable to chromatography, so they
usually do separate out pretty well. So you don’t have terribly
complex samples but if you have a pretty good chromatography
set up you might be able to separate out your mixture into say
1,000 or 2,000 fractions and if you’ve got 100,000 or a million
infractions that means that you’ve got quite a few components
coming out at any particular time and you still need to have high
resolution to be able to deal with the complexity of the sample.
For proteomics, the approach is to try to run the experiment and
take advantage of the high performance of the mass spectrometer
for selection and tandem mass spec. They have to run the mass
spectrometer really, really fast and try to select as many peaks
as possible in every time slice in order to fragment them and
get sequence information. This approach does work, and we do
that too, but we have the added advantage of having very high
resolution meaning we can separate out the mixtures at the same
time. Which helps us process this data. Then there’s all the other
elements such as glycomics and genomics that are also amenable
to this high-performance mass spectrometry in different
contexts.
Q: Your team’s mission is to “develop new FTICR
mass spectrometry instruments with unique
capabilities.” Could you elaborate on this?
A: Development is most of what we do. We work on developing
advanced FTICR mass spectrometry, more specifically, we
develop new tools using the high-resolution capabilities of the
FTICR.
I’m currently working on building a web app that I intend
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 The Diverse Applications of Mass Spectrometry
to use to teach people how to calculate isotope distribution.
Isotope distributions are not actually very complicated, but
it really is important to get them right if you’re going to be
comparing them to real data. A lot of web applications that
you find make some pretty simple assumptions that are often
fine – but can sometimes cause problems. We have to calculate
isotopic distributions for all metals which is not always easy.
We also work on different ways of fragmenting molecules. In
the mass spectrometer you will typically measure your mass
spectrum and get a number of peaks for peptides. You’ll then
select one peak using some of the voltages and wave forms that
we have in the mass spectrometer, smash it using a tandem mass
spectrometry fragmentation method and then measure the
masses that you’ve created – which will presumably allow you
to get some structural information for peptides. This method
usually enables the partial (or sometimes full) sequencing of the
peptides. So there’s a lot of tricks to sequencing using tandem
mass spectrometry.
How do you break the molecules? Most of the time, we collide
them with background gas and break them solely by collisions.
We are also working on doing photo dissociation which involves
breaking molecules with UV and IR light, and we also do
reactions with electrons. There are half a dozen different types
of reactions you can perform with electrons that causes the
molecules to break in different ways. Each reaction tends to
result in slightly different information, so when you combine
several of these reactions together you can get a very complete
picture of sequence information which is very useful. We’ve
actually developed a new fragmentation technique, called two
dimensional mass spectrometry, which is very exciting. We’ve
got a few papers out on it now and we’re about to submit three or
four more.
Two-dimensional mass spectrometry technology is well
established. The concepts and methodologies were worked back
in the late 80s, but it required too much computational power
for the time, we needed modern computers to be able to start to
successfully process the data. When it comes to two-dimensional
mass spectrometry, you may have one spectrum that equates
to 200 Gb of data – your average computer can’t process that, it
requires cluster computers, housed here in the university. This
enhanced computing power allows us to get it down into a format
that we can use. The technology finally enabled us to use this
method even though it’s been sitting there for close to 30 years in
the literature! It allows us to fragment the molecules.
In two-dimensional mass spectrometry you don’t have to
isolate the molecules. What you do instead is you modulate all
of the molecules in and out of a fragmentation zone and you
basically code each pre-cursor ion with its own modulation
frequency and because that is dragging it back and forth to the
fragmentation zone the fragments end up carrying the same
Technology Networks 2018
modulation frequency as the pre-cursor. This means that you
can fragment all the molecules at the same time without doing
any isolations and it greatly speeds up the whole process because
you don’t have to do them one by one, you just do them all
together. It speeds it up and it also gives you much more detail in
the information because what inevitably happens with complex
samples is you can fragment the first 5 or 10 and then you run out
of time to be able to do the next bit so it’s basically a much faster
way of doing the whole experiment. It will end up giving us a lot
more information about structures and molecules in complex
mixtures. It’s an ongoing development project though so stay
tuned on that one I guess!
Q: What projects are you currently working on?
A: Two-dimensional mass spectrometry; that’s a big
development project for us going forward and we’re working on
it in pretty much every avenue that we can think to apply it. We
have recently had two scientific papers accepted that outline the
technology and its use for the analysis of whole proteins rather
than the analysis of smaller peptides that you would typically
create by digesting the proteins – top-down proteomics. Of
course, we’re using it on proteomics and polymers and we are
also making good progress on the use of two-dimensional mass
spectrometry for glycomics. There are a few application areas
that are particularly interesting.
The first project to mention is in collaboration with a medical
school in Beijing. The researchers have scorpion venom and
scorpion venom extracts that they are using that have particular
anti-coagulant activities. The idea is to try to figure out what
is it in the structures of these scorpion venom extracts that is
responsible for the anti-coagulant activity. It turns out that most
of the molecules in the extract are proteins, not very big ones,
pretty small, under 10,000 daltons, but they’re tied up in really
tight balls with many disulphide bonds, presumably so that
they’ll survive when they’re out in the environment and being
in this conformation prevents them from immediately being
chopped up by proteases and allows them to maintain their
stability. But, first of all, you need to try to figure out what these
molecules are. The scorpions aren’t sequenced so we don’t have a
database to search against so we have to sequence them ‘de novo’,
using just the mass spectrometry data - which is much harder.
We also must try to figure out exactly where those disulfide
bonds are, you tend to just scramble where the disulfide bonds
are (if you even keep that information) so it’s a challenge to try
to figure out exactly what these venoms are and how they work.
That’s a fun project which is making some nice progress at the
moment.
We are also working on analysis of amyloid plaques in
Alzheimer’s disease patients, specifically trying to figure out
what impact metallic particles have on Alzheimer’s disease.
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 The Diverse Applications of Mass Spectrometry

There has been a lot of discussion about aluminium and iron in

your diet or that you pick up from environmental pollution and
how this might impact Alzheimer’s disease. I have a collaborator
who works on that and our part of the project is to try to figure
out what modifications we see in the Alzheimer’s disease
proteins, the amyloid beta protein, as a function of these metallic
nanoparticles. They also then go off to the synchrotrons and try
to figure out exactly what the chemical state of the metal is in
that particular nanoparticle. Is it actually a metal or is it a metal
that is bound to something else? It is an interesting problem from
a lot of different directions, technologically as well as in terms of
medicine.

We also have a big project on antibodies. This is a collaboration

project with a biopharmaceutical company, who manufacture
and use a lot of different antibodies. They’re extremely interested
in all the post-translational modifications that feature on them. If
these antibodies are being produced for pharmaceutical purposes
they want to ensure that the antibodies remain stable over time
and that they aren’t creating some kind of side reaction that
then causes problems with the activity of the drug. Therefore,
there is a lot of work currently focused on post-translational
modifications of antibodies.

Peter B. O’Connor was speaking to Laura Elizabeth Mason, Science Writer

for Technology Networks.

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